牛病毒性腹泻病毒E^rns-E2融合蛋白在果蝇S2细胞中的表达与鉴定 |
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引用本文: | 孙宇,史利军,孙卫华,章金刚. 牛病毒性腹泻病毒E^rns-E2融合蛋白在果蝇S2细胞中的表达与鉴定[J]. 生物技术通讯, 2008, 19(6): 862-865 |
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作者姓名: | 孙宇 史利军 孙卫华 章金刚 |
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作者单位: | [1]军事医学科学院野战输血研究所,北京100850 [2]解放军第305医院,北京100017 |
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摘 要: | 目的:利用果蝇S2细胞表达牛病毒性腹泻病毒(BVDV)Erns-E2融合蛋白,并对其抗体结合能力进行鉴定。方法:用RT-PCR方法扩增BVDV NADL株Erns和E2蛋白的编码基因,利用(G4-S)3柔性15肽基因将扩增的2个基因连接,再与昆虫表达载体pMT/BiP/V5-His连接构建重组表达载体pMT/BiP/V5-His-Erns-E2,将后者与筛选质粒pCoBlast共转染果蝇S2细胞后表达Erns-E2融合蛋白,并对表达产物进行鉴定。结果:SDS-PAGE结果表明,融合蛋白相对分子质量为76800;Western blotting检测表明,该融合蛋白具有与BVDV抗体良好的结合能力。结论:BVDV的Erns-E2融合蛋白能在果蝇S2细胞中进行表达;经鉴定,表达产物具有良好的抗体结合能力,可用于抗原检测。
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关 键 词: | 牛病毒性腹泻病毒 E^rns蛋白 E2蛋白 融合表达 |
Expression and Identification of the Fusion Protein E^rns-E2 of Bovine Viral Diarrhea Virus in Drosophila $2 Cell |
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Affiliation: | SUN Yu, SHI Li-Jun, SUN Wei-Hua, ZHANG Jin-Gan( 1. Institute of Transfusion Medicine, Academy of Military Medical Sciences, Beijing 100850; 2. The 305 Military Hospital, Beijing 100017; China) |
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Abstract: | Objective: To expressE^rns-E2 fusion protein of bovine viral diarrhea virus(BVDV) by Drosophila S2 protein expressed system and to detect the antigenieity of the fusion protein. Methods: The genes endoding E^rns and E2 of BVDV NADL strain were amplified by RT-PCR, and were ligated by (G4-S)3 gene as fusion gene, then was cloned into pMT/BiP/V5-His vector for constructing the recombinant expression vector pMT/BiP/V5-His-E^rns-E2 Expression vector pMT/BiP/V5-His-E~-E2 and screen vector pCoBlast were cotransfected into s2 cells to generate fusion protein. The anti- genieity of expressed protein was identified by Western blotting. Results: A specific and simple band at about 76.8 kDa was showed by SDS-PAGE and the protein was proved to have good antibody binding capacity by Western blotting analysis. Conclusion: This study indicated that E^rns-E2 fusion gene of BVDV NADL strain can be effective translated in S2 cells as respected, and theE^rns-E2 fusion protein can be used as an envelope antigen for BVDV antibody detection. |
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Keywords: | bovine viral diarrhea virus E^rns protein E2 protein fusion expression |
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